Biocidal compositions and use thereof containing a synergistic mixture of 2-bromo-2-nitropropane-1,3-diol and tri-n-butyl tetradecyl phosphonium chloride

ABSTRACT

Bactericidal composition and method for inhibiting and controlling the growth of the capsulated, facultative bacterium, Klebsiella pneumoniae, are disclosed. The composition comprises an amount, effective for the intended purpose, of 2-bromo-2-nitropropane-1,3-diol (BNPD) and &#34;Belclene 350.&#34; The active biocidal component of the &#34;Belclene 350&#34; product is tri n-butyl tetradecyl phosphonium chloride. The method comprises administering between about 0.1 to about 200 parts of this composition (i.e., BNPD and &#34;Belclene 350&#34; product) (based on one million parts of the desired aqueous system) to the particular water containing system for which treatment is desired.

BACKGROUND OF THE INVENTION

The formation of slimes by microorganisms is a problem that isencountered in many aqueous systems. For example, the problem is notonly found in natural waters such as lagoons, lakes, ponds, etc. andconfined waters as in pools, but also in such industrial systems ascooling water systems, air washer systems and pulp and paper millsystems. All possess conditions which are conducive to the growth andreproduction of slime-forming microorganisms. In both once-through andrecirculating cooling systems, for example, which employ largequantities of water as a cooling medium, the formation of slime bymicroorganisms is an extensive and constant problem.

Airborne organisms are readily entrained in the water from coolingtowers and find this warm medium an ideal environment for growth andmultiplication. Aerobic and heliotropic organisms flourish on the towerproper while other organisms colonize and grow in such areas as thetower sump and the piping and passages of the cooling system. The slimeformation not only aids in the deterioration of the tower structure inthe case of wooden towers, but also, by its deposition on metalsurfaces, promotes corrosion, In addition, slime carried through thecooling system plugs and fouls lines, valves, strainers, etc. anddeposits on heat exchange surfaces. In the latter case, the impedance ofheat transfer can greatly reduce the efficiency of the cooling system.

In pulp and paper mill systems, slime formed by microorganisms iscommonly encountered and causes fouling, plugging, or corrosion thereof.The slime also becomes entrained in the paper produced to causebreakouts on the paper machines, with consequent work stoppages and theloss of production time, and/or is responsible for unsightly blemishesin the final product, which result in rejects and wasted output.

The previously discussed problems have resulted in the extensiveutilization of biocides in cooling water and pulp and paper millsystems. Materials which have enjoyed widespread use in suchapplications include chlorine, chlorinated phenols, organo-bromines, andvarious organo-sulfur compounds. All of these compounds are generallyuseful for this purpose but each is attended by a variety ofimpediments. For example, chlorination is limited both by its specifictoxicity for slime-forming organisms at economic levels and by thetendency of chlorine to react, which results in the expenditure of thechlorine before its full biocidal function is achieved. Other biocidesare attended by odor problems and hazards in respect to storage, use orhandling which limit their utility. To date, no one compound or type ofcompound has achieved a clearly established predominance in respect tothe applications discussed. Likewise, lagoons, ponds, lakes, and evenpools, either used for pleasure purposes or used for industrial purposesfor the disposal and storage of industrial wastes, become, during thewarm weather, beseiged by slime due to microorganism growth andreproduction. In the case of the recreational areas the problem ofinfection is obvious. In the case of industrial storage or disposal ofindustrial materials, the microorganisms cause additional problems whichmust be eliminated prior to the material's use or disposal of the waste.

Naturally, economy is a major consideration in respect to all of thesebiocides. Such economic considerations attach to both the cost of thebiocide and the expense of its application. The cost performance indexof any biocide is derived from the basic cost of the material, itseffectiveness per unit of weight, the duration of its biocidal orbiostatic effect in the system treated, and the same and frequency ofits addition to the system treated. To date, none of the commerciallyavailable biocides has exhibited a prolonged biocidal effect. Instead,their effectiveness is rapidly reduced as the result of exposure tophysical conditions such as temperature, association with ingredientscontained by the system toward which they exhibit an affinity orsubstantivity, etc., with a resultant restriction or elimination oftheir biocidal effectiveness, or by dilution.

As a consequence, the use of such biocides involves their continuous orfrequent addition to systems to be treated and their addition to aplurality of points or zones in the systems to be treated. Accordingly,the cost of the biocide and the labor cost of such means of applying itare considerable. In other instances, the difficulty of access to thezone in which slime formation is experienced precludes the effective useof a biocide. For example, if in a particular system there is no accessto an area at which slime formation occurs the biocide can only beapplied at a point which is upstream in the flow system. However, thephysical or chemical conditions, e.g., chemical reactivity, thermaldegradation, etc. which exist between the point at which the biocide maybe added to the system and the point at which its biocidal effect isdesired render the effective use of a biocide impossible.

Similarly, in a system experiencing relatively slow flow, such as apaper mill, if a biocide is added at the beginning of the system, itsbiocidal effect may be completely dissipated before it has reached allof the points at which this effect is desired or required. As aconsequence, the biocide must be added at a plurality of points, andeven then a diminishing biocidal effect will be experienced between onepoint of addition to the system and the next point downstream at whichthe biocides may be added. In addition to the increased cost ofutilizing and maintaining plural feed points, gross ineconomies inrespect to the cost of the biocide are experienced. Specifically, ateach point of addition, an excess of the biocide is added to the systemin order to compensate for that portion of the biocide which will beexpended in reacting with other constituents present in the system orexperience physical changes which impair its biocidal activity.

SUMMARY OF THE INVENTION

The biocidal compositions of the present invention comprise, as activeingredients, (1) 2-bromo-2-nitropropane-1,3-diol (hereinafter "BNPD")and (2) a tetraalkyl phosphonium halide compound (TPH).

PRIOR ART

BNPD is sold by The Boots Company, Ltd., Nottingham, England, as anindustrial water treatment antibacterial agent.

The specific TPH herein preferred for use is tri n-butyl tetradecylphosphonium chloride. This particular tetraalkyl phosphonium halide iscommercially available from Ciba-Geigy under the trademark "Belclene350" and is touted as a "broad spectrum biocide specifically developedfor the control of microbiological fouling in cooling water systems."

DETAILED DESCRIPTION

Surprisingly, the present inventors have found that a compositioncomprising BNPD and TPH is especially efficacious in controlling thegrowth of bacterial microbes, specifically the Klebsiella pneumoniaespecies. This particular species is a member of the capsulated,facultative class of bacteria and is generally present in air, water andsoil. These bacteria continually contaminate open cooling systems andpulping and papermaking systems and are among the most common slimeformers. This slime may be viewed as being a mass of agglomerated cellsstuck together by the cementing action of the gelatinous polysaccharideor proteinaceous secretions around each cell. The slimy mass entrapsother debris, restricts water flow and heat transfer, and may serve as asite for corrosion.

The fact that the Klebsiella species used in the tests is a facultativespecies is important as, by definition, such bacteria may thrive undereither aerobic or anaerobic conditions. Accordingly, by reason ofdemonstrated efficacy in the growth inhibition of this particularspecies, one can expect similar growth inhibition attributes when otheraerobic or anaerobic bacterial species are encountered.

As above noted, BNPD is available from The Boots Company, Ltd. and issold under the trademarks "Myacide AS" or "Bronopol." It is a white freeflowing crystalline solid that is readily soluble in cold water. Theproduct is from about 95-100% pure.

The physical properties of "Belclene 350" are reported as being:

    ______________________________________                                        Appearance        Clear, colorless liquid                                     Specific gravity at 20° C.                                                               0.96                                                        pH                7.0-8.0                                                     Boiling point     100° C.                                              Freezing point    -8 to -10° C.                                        Viscosity         50-80 cP                                                    Odor              Slight                                                      Solubility:                                                                   Water             Completely miscible in all                                                    proportions                                                 Methanol          Greater than 50%                                            Isopropanol       Greater than 50%                                            Ethylene glycol   Greater than 50%                                            ______________________________________                                    

In accordance with the present invention, the combined BNPD:"Belclene350" treatment may be added to the desired aqueous system in need ofbiocidal treatment, in an amount of from about 0.1 to about 200 parts ofthe combined treatment to one million parts (by weight) of the aqueousmedium. Preferably, about 5 to about 50 parts of the combined treatmentper one million parts (by weight) of the aqueous medium is added.

The combined treatment is added, for example, to cooling water systems,paper and pulp mill systems, pools, ponds, lagoons, lakes, etc., tocontrol the formation of bacterial microorganisms, which may becontained by, or which may become entrained in, the system to betreated. It has been found that the BNPD/"Belclene 350" compositions andmethods of utilization of the treatment are efficacious in controllingthe facultative bacterium, Klebsiella pneumoniae, which may populatethese systems. It is thought that the combined treatment composition andmethod of the present invention will also be efficacious in inhibitingand controlling all types of aerobic and anaerobic bacteria.

Surprisingly, it has been found that when the BNPD/"Belclene 350"ingredients are mixed, in certain instances, the resulting mixturespossess a higher degree of bacterial activity than that of theindividual ingredients comprising the mixture. Accordingly, it ispossible to produce a highly efficacious bactericide. Because of theenhanced activity of the mixture, the total quantity of the bacterialtreatment may be reduced. In addition, the high degree of bacterialeffectiveness which is provided by each of the ingredients may beexploited without use of higher concentrations of each.

The following experimental data were developed. It is to be rememberedthat the following examples are to be regarded solely as beingillustrative, and not as restricting the scope of the invention.

DESCRIPTION OF PREFERRED EMBODIMENT

BNPD and "Belclene 350" were added in varying ratios and over a widerange of concentrations to a liquid nutrient medium which wassubsequently inoculated with a standard volume of a suspension of thefacultative bacterium Klebsiella pneumoniae. Growth was measured bydetermining the amount of radioactivity accumulated by the cells when ¹⁴C-glucose was added as the sole source of carbon in the nutrient medium.The effect of the biocide chemicals, alone and in combination, is toreduce the rate and amount of ¹⁴ C incorporation into the cells duringincubation, as compared to controls not treated with the chemicals.Additions of the biocides, alone and in varying combinations andconcentrations, were made according to the accepted "checkerboard"technique described by M. T. Kelley and J. M. Matsen, AntimicrobialAgents and Chemotherapy. 9: 440, (1976). Following a two hourincubation, the amount of radioactivity incorporated in the cells wasdetermined by counting (¹⁴ C liquid scintillation procedures) for alltreated and untreated samples. The percent reduction of each treatedsample was calculated from the relationship: ##EQU1##

Plotting the % reduction of ¹⁴ C level against the concentration of eachbiocide acting alone results in a dose-response curve, from which thebiocide dose necessary to achieve any given % reduction can beinterpolated.

Synergism was determined by the method of calculation described by F. C.Kull, P. C. Eisman, H. D. Sylwestrowicz and R. L. Mayer, AppliedMicrobiology 9, 538, (1961) using the relationship: ##EQU2## where:Q_(a) =quantity of compound A, acting alone, producing an end point

Q_(b) =quantity of compound B, acting alone, producing an end point

Q_(A) =quantity of compound A in mixture, producing an end point

Q_(B) =quantity of compound B in mixture, producing an end point

The end point used in the calculations is the % reduction caused by eachmixture of A and B. Q_(A) and Q_(B) are the individual concentrations inthe A/B mixture causing a given % reduction. Q_(a) and Q_(b) aredetermined by interpolation from the respective dose-response curves ofA and B as those concentrations of A and B acting alone which producethe same % reduction as each specific mixture produced.

Dose-response curves for each active acting alone were determined bylinear regression analysis of the dose-response data. Data were fittedto a curve represented by the equation shown with each data set. Afterlinearizing the data, the contributions of each biocide component in thebiocide mixtures to the inhibition of radioisotope uptake weredetermined by interpolation with the dose response curve of therespective biocide. If, for example, quantities of Q_(A) plus Q_(B) aresufficient to give a 50% reduction in ¹⁴ C content, Q_(a) and Q_(b) arethose quantities of A or B acting alone, respectively, found to give 50%reduction in ¹⁴ C content. A synergism index (SI) is calculated for eachcombination of A and B.

Where the SI is <1, synergism exists. Where the SI=1, additivity exists.Where SI>1, antagonism exists.

The data in the following tables come from treating Klebsiellapneumoniae, a common nuisance bacterial type found in industrial coolingwaters and in pulping and paper making systems, with varying ratios andconcentrations of BNPD and "Belclene 350." Shown for each combination isthe % reduction of ¹⁴ C content, the calculated SI, and the weight ratioof BNPD to "Belclene 350."

                  TABLE I                                                         ______________________________________                                        BNPD vs "Belclene 350"                                                                 ppm**     ratio                                                      ppm      "Belclene BNPD:Bel-                                                  BNPD     350"      clene 350   % I  SI                                        ______________________________________                                        2.5       0        100:0        0                                             5         0        100:0        0                                             10        0        100:0        0                                             20        0        100:0       14                                             40        0        100:0       78                                             80        0        100:0       95                                             0        20         0:100      12                                             0        25         0:100      18                                             0        40         0:100      47                                             0        50         0:100      62                                             0        80         0:100      90                                             0        100        0:100      94                                             2.5      100        1:40       97   1.11                                      5        100        1:20       97   1.14                                      10       100        1:10       96   1.22                                      20       100       1:5         97   1.32                                      40       100         1:2.5     99   1.59                                      80       100         1:1.25    99   2.12                                      2.5      80         1:32       93   0.93*                                     5        80         1:16       96   0.94*                                     10       80        1:8         95   1.02                                      20       80        1:4         98   1.13                                      40       80        1:2         99   1.38                                      80       80        1:1         99   1.91                                      2.5      50         1:20       79   0.70*                                     5        50         1:10       74   0.79*                                     10       50        1:5         86   0.75*                                     20       50          1:2.5     97   0.81*                                     40       50          1:1.25    98   1.09                                      80       50        1.6:1       99   1.59                                      2.5      40         1:16       57   1.16                                      5        40        1:8         64   0.75*                                     10       40        1:4         81   0.67*                                     20       40        1:2         94   0.72*                                     40       40        1:1         97   0.98                                      80       40        2:1         99   1.49                                      2.5      25         1:10       25   1.11                                      5        25        1:5         32   0.97                                      10       25          1:2.5     61   0.63*                                     20       25          1:1.25    84   0.62*                                     40       25        1.6:1       96   0.82*                                     80       25        3.2:1       99   1.33                                      2.5      20        1:8         11   1.89                                      5        20        1:4         21   1.17                                      10       20        1:2         48   0.69*                                     20       20        1:1         86   0.54*                                     40       20        2:1         96   0.77*                                     80       20        4:1         99   1.28                                      ______________________________________                                    

                  TABLE II                                                        ______________________________________                                        BNPD vs "Belclene 350"                                                                 ppm**     ratio                                                      ppm      "Belclene BNPD:Bel-                                                  BNPD     350"      clene 350   % I  SI                                        ______________________________________                                        2.5       0        100:0        0                                             5         0        100:0        0                                             10        0        100:0        0                                             20        0        100:0       18                                             40        0        100:0       76                                             80        0        100:0       94                                             0        20         0:100      19                                             0        25         0:100      30                                             0        40         0:100      43                                             0        50         0:100      63                                             0        80         0:100      77                                             0        100        0:100      85                                             2.5      100        1:40       91   1.04                                      5        100        1:20       91   1.07                                      10       100        1:10       93   1.12                                      20       100       1:5         97   1.20                                      40       100         1:2.5     98   1.45                                      80       100         1:1.25    99   1.74                                      2.5      80         1:32       73   1.07                                      5        80         1:16       85   0.95*                                     10       80        1:8         88   0.98                                      20       80        1:4         95   1.04                                      40       80        1:2         98   1.27                                      80       80        1:1         99   1.56                                      2.5      50         1:20       65   0.78*                                     5        50         1:10       71   0.76*                                     10       50        1:5         79   0.75*                                     20       50          1:2.5     94   0.76*                                     40       50           1:1.125  97   1.00                                      80       50        1.6:1       99   1.29                                      2.5      40         1:16       51   0.86*                                     5        40        1:8         53   0.87*                                     10       40        1:4         71   0.72*                                     20       40        1:2         89   0.70*                                     40       40        1:1         97   0.91*                                     80       40        2:1         98   1.43                                      2.5      25         1:10       27   1.36                                      5        25        1:5         43   0.77*                                     10       25          1:2.5     53   0.71*                                     20       25          1:1.25    88   0.55*                                     40       25        1.6:1       96   0.78*                                     80       25        3.2:1       98   1.30                                      2.5      20        1:8         16   3.02                                      5        20        1:4         27   1.28                                      10       20        1:2         43   0.79*                                     20       20        1:1         79   0.56*                                     40       20        2:1         96   0.73*                                     80       20        4:1         98   1.25                                      ______________________________________                                         *In Tables I and II, asterisks in the S.I. column indicate synergistic        combinations in accordance with the Kull et al. method supra.                 **The double asterisk indicates that the "Belclene 350" product tested        consisted of 50% (wt.) active biocidal component.                        

In Tables I and II, differences seen between the replicates are due tonormal experimental variance.

In accordance with Tables I-II supra., unexpected results occurred morefrequently within the product ratios of BNPD:"Belclene 350" product offrom about 2:1 to 1:32 with a product ratio of BNPD:"Belclene" of about2:1 to 1:20 being even more preferred. Since the "Belclene 350" productcontains about 50% active biocidal component (i.e., tri n-butyltetradecyl phosphonium chloride), when based on the active biocidalcomponents, unexpected results appear more frequently within the rangeof BNPD:active component of "Belclene 350" of about 4:1 to 1:16 with anactive component range of BNPD:active component of "Belclene 350" of 4:1to 1:10 being even more preferred. At present, it is most preferred thatany commercial product embodying the invention comprises a weight ratioof active component of about 2:1 BNPD:tri n-butyl tetradecyl phosphoniumchloride.

While we have shown and described herein certain embodiments of thepresent invention, it is intended that there be covered as well anychange or modification therein which may be made without departing fromthe spirit and scope of the invention.

We claim:
 1. A bacterial inhibiting composition comprising a synergisticaqueous mixture of (a) 2-bromo-2-nitropropane-1,3-diol (BNPD) and (b)tri n-butyl tetradecyl phosphonium chloride, the weight ratio of (a):(b)being from about 2:1 to about 1:16.
 2. The composition as recited inclaim 1 wherein the weight ratio of (a) to (b) is about 2:1.
 3. A methodfor controlling the growth of Klebsiella pneumoniae bacteria in anaqueous system which comprises adding to said system from about 0.1 toabout 200 parts per weight of a synergistic composition per one millionparts per weight of said aqueous system, said composition comprising (a)2-bromo-2-nitropropane-1,3-diol (BNPD), and (b) tri n-butyl tetradecylphosphonium chloride, the weight ratio of (a):(b) being from about 2:1to about 1:16.
 4. The method as recited in claim 3 wherein (b) is in theform of an aqueous composition comprising about 50% active biocidalingredient.
 5. The method as recited in claim 3 wherein the weight ratioof (a):(b) is about 2:1.
 6. The method as defined in claim 3 whereinsaid composition is added to said system in an amount of from about 5 toabout 50 parts per million of said aqueous system.
 7. The method asdefined in claim 3 wherein said aqueous system comprises a cooling watersystem.
 8. The method as defined in claim 3 wherein said aqueous systemcomprises a pulping and papermaking system.